Synthesis of novel tubulin polymerization inhibitors: benzoylphenylurea (bpu) sulfur analogs

ABSTRACT

A novel series of BPU analogues were synthesized and evaluated for antitumor activity. In particular, BPU sulfur analogues 6n and 7d were shown to possess up to 10-fold increased potency, when compared to compound 1, against cancer cell lines. 6n was more effective than compound 1 in causing apoptosis of MCF-7 cells. When compared to other drugs with a similar mechanism of action, 6n retained significant ability to inhibit tubulin assembly, with an IC50 of 2.1 μM.

CROSS-REFERENCES TO RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.11/673,519 which is a continuation of PCT/US2006/014449, filed Apr. 18,2006 and published as WO 2006/113650, which claims benefit of priorityfrom U.S. Provisional Patent Application Nos. 60/672,469, filed Apr. 18,2005; 60/696,672 filed Jul. 5, 2005; and 60/720,755 filed Sep. 27, 2005.All five applications are incorporated herein by reference in theirentireties for all purposes.

STATEMENT AS TO RIGHTS TO INVENTIONS MADE UNDER FEDERALLY SPONSOREDRESEARCH OR DEVELOPMENT

This study was supported in-part by National Cancer Institute Contract#N01-CO-12400 and all joint inventors have assigned their rights toJohns Hopkins University.

BACKGROUND OF THE INVENTION

1. Field of Invention

The present invention relates to design and synthesis of novelbenzoylphenylurea (BPU) sulfur analogs with increased potency andenhanced water solubility. More specifically, the present invention isdirected toward the synthesis of sulfur, hydroximic acid, and boronicacid analogs of BPU.

2. Brief Description of Art

The lack of selectivity of many cancer agents and the occurrence ofintrinsic or acquired resistance of tumors to chemotherapy has beenmajor obstacles in the treatment of cancer. Microtubules, which are keycomponents of the cell, play an important role in a variety of cellularprocess, including mitosis and cell division. See, Downing, K. H., etal, Curr. Opiti. Struct. Biol., 5:785-791 (1998) and Sorger, P. K., etal., Curr. Opin. Cell Biol., 9:807-814 (1997). Antimitotic agents can bedivided into two major classes: (1) microtubule stabilizers such aspaclitaxel and docetaxel, which prevent the dcpolymerization of tubulin(Jordan, A., et al., Med. Res. Rev., 18:259-296 (1998)); and (2) Vincaalkaloids (e.g., vincristine, vinblastine, and vinorelbine) andcolchicines, which inhibit the polymerization of tubulin (Jordan, A., etal, Med. Res. Rev., 18:259-296 (1998)). Although some of these agentshave been used as antiprohferative agents in the treatment of humanmalignancies, they suffer from drug resistance mediated through theexpression of efflux pumps. See, Lehnert, M., Eur. J. Cancer,32A:912-920 (1996) and Germann, U. A., Eur. J. Cancer, 32A:927-944(1996).

Urea and thiourea derivatives have been used for the treatment of a widerange of solid tumors, Easmon, J. et al.; J. Med. Chem., 44:2164-2171(2001). Urea-based prodrugs have been reported as candidates formelanocyte-directed enzyme prodrug therapy (MDEPT), in which theyrelease the drug upon exposure to tyrosinase. See, Jordan, A. M. et al.,Bioorg. Med. Chem., 10:2625-2633 (2002). Benzoylphenylurea (BPU)compounds were originally developed as insecticides, and their antitumoractivity was found during random screening (Okada, H., et al., Chem.Pharm. Bull., 39:2308-2315 (1991)). Various analogues of BPU weresynthesized, and their cytotoxic activity was examined to establishstructure-activity relationships (Okada, H., et al., Chem. Pharm. Bull.,39:2308-2315 (1991)). To improve physicochemical properties, organic andwater soluble BPU derivatives were developed. See, Okada, H., et al.,Chem. Pharm. Bull., 42:57-61 (1994); and Okada, H., et al., Chem. Pharm.Bull., 47:430-433 (1999). Six of these analogues were screened at theNCl for their cytotoxicity against various cancer cell lines. Theyexhibited potent antitumor activity in vitro against several cancer celllines, as well in vivo against several tumor models. These compoundsalso have been reported to be effective inhibitors of tubulinpolymerization (Holligshead, M. G., et al., Proc. Am. Assoc. CancerRes., 39:164 (1979)). One of these agents, compound 1 (NSC-639829),

(Holligshead, M. G., et al., Proc. Am. Assoc. Cancer Res., 39:164(1979)) is currently being evaluated in Phase T clinical trials inpatients with refractory metastatic cancer. See, Messersmith, W. A., etal., Proc. Am. Soc. Clin. Oncol., 22:203 (2003); and Edelman, M. J., etal., Proc. Am. Soc. Clin. Oncol., 22:137 (2003). We have previouslyreported the synthesis and antitumor evaluation of a set of BPUanalogues (Gurulingappa, H., et al., Bioorg. Med. Chem. Lett.,14:2213-2216 (2004)).

There is still a need to improve physicochemical properties, organic andwater soluble BPU derivatives. To that end the present inventiondiscloses a novel series of synthesized derivatives of BPU by replacingthe urea moiety with thiourea and the ether linkage with sulfide,sulfoxide, or sulfone groups. The activity of such novel agents, aresignificantly more toxic to cancer cells in culture.

BRIEF SUMMARY OF THE INVENTION

In one aspect, the present invention provides an antiproliferative,water soluble compound with high bioavailability having the generalformula (I)

where A, B, and C are independently substituted alkyl, cycloalkyl, aryl,heterocycle, or substituted phenyl fused with a saturated or unsaturated4- to 6-membered rings optionally containing one to three heteroatomsindependently selected from N, O, and S; ‘X’ is oxygen or sulfur; ‘Y’ issulfur, sulfoxide or sulfone; X₁ is CO or hydrogen; R₅ is hydrogen, oralkyl or aryl group; X₂ is hydrogen or SR₆, wherein R₆ is a substitutedalkyl, cycloalkyl, aryl, heterocycle, or substituted phenyl fused with asaturated or unsaturated 4- to 6-membered rings optionally containingone to three heteroatoms independently selected from N, O, and S.

In another aspect, the present invention provides pharmaceuticalcompositions. The pharmaceutical composition includes a pharmaceuticallyacceptable excipient and a compound having the formula (I):

where A, B, and C are independently substituted alkyl, cycloalkyl, aryl,heterocycle, or substituted phenyl fused with a saturated or unsaturated4- to 6-membered rings optionally containing one to three heteroatomsindependently selected from N, O, and S; ‘X’ is oxygen or sulfur; ‘Y’ issulfur, sulfoxide or sulfone; X₁ is CO or hydrogen; R₅ is hydrogen, oralkyl or aryl group; X₂ is hydrogen or SR₆, wherein R₆ is a substitutedalkyl, cycloalkyl, aryl, heterocycle, or substituted phenyl fused with asaturated or unsaturated 4- to 6-membered rings optionally containingone to three heteroatoms independently selected from N, O, and S.

BRIEF DESCRIPTION OF THE DRAWINGS

The accompanying drawings, which are incorporated in and form a part ofthe specification, illustrate the preferred embodiments of the presentinvention, and together with the description serve to explain theprinciples of the invention.

In the drawings, FIGS. 8-16, the cytotoxic effects of the test compoundson seven pancreatic cell lines was measured. The horizontal axis depictsvarious dilutions of the test compounds that were exposed to pancreaticcell lines. The vertical axis (cell number) depicts the number ofpancreatic cells present after exposure to a specific concentration ofthe tested compound as compared to the cell number at time zero.

In the drawings, FIGS. 17-34, the growth inhibition effects of the testcompounds on prostate cancer cell lines LAPC-4 (FIGS. 10-15); CWR22R(FIGS. 16-21); and LnCaP (FIGS. 22-27) are measured. The horizontal axisdepicts the days after exposure on which the assays were performed. Thevertical axis (cell number) depicts the number of pancreatic cellspresent after exposure to a specific concentration of the testedcompound as compared to the cell number at time zero.

In the Drawings:

FIG. 1 schematically depicts the synthesis scheme for compounds of thepresent invention.

FIG. 2 schematically depicts the synthesis of the Ring-A anilinemodified amino acids prodrugs.

FIG. 3 schematically depicts the synthesis scheme for ring-A anilinemodified carbamylosulfenyl derivative of BPU sulfur analogs.

FIG. 4 schematically depicts the synthesis scheme for compounds 4a and4b of the present invention.

FIG. 5 schematically depicts the synthesis scheme for compounds of thepresent invention.

FIG. 6 schematically depicts the synthesis scheme for both ring-a andurea modified prodrugs of the present invention.

FIG. 7 is a bar graph demonstrating the apoptosis of MCF-7 cells bycompounds 6n and 1.

FIG. 8 depicts the dose response curves generated by exposing sevenpancreatic cell lines to various concentrations of compound 1 as acontrol.

FIG. 9 depicts the dose response curves generated by exposing sevenpancreatic cell lines to various concentrations of compound 6n of thepresent invention.

FIG. 10 depicts the dose response curves generated by exposing sevenpancreatic cell lines to various concentrations of compound 7d of thepresent invention.

FIG. 11 depicts the dose response curves generated by exposing sevenpancreatic cell lines to various concentrations of compound 6d of thepresent invention.

FIG. 12 depicts the dose response curves generated by exposing sevenpancreatic cell lines to various concentrations of compound 6h of thepresent invention.

FIG. 13 depicts the dose response curves generated by exposing sevenpancreatic cell lines to various concentrations of compound 7b of thepresent invention.

FIG. 14 depicts the dose response curves generated by exposing sevenpancreatic cell lines to various concentrations of compound 6g of thepresent invention.

FIG. 15 depicts the dose response curves generated by exposing sevenpancreatic cell lines to various concentrations of compound 8g of thepresent invention.

FIG. 16 depicts the dose response curves generated by exposing sevenpancreatic cell lines to various concentrations of compound 8h of thepresent invention.

FIG. 17 depicts the dose response curves generated by exposing variousconcentrations of compound 1 of the present invention to human prostatecancer cell line LAPC-4 versus a control using only a solvent.

FIG. 18 depicts the dose response curves generated by exposing variousconcentrations of compound 6l of the present invention to human prostatecancer cell line LAPC-4 versus a control using only a solvent.

FIG. 19 depicts the dose response curves generated by exposing variousconcentrations of compound 6h of the present invention to human prostatecancer cell line LAPC-4 versus a control using only a solvent.

FIG. 20 depicts the dose response curves generated by exposing variousconcentrations of compound 6n of the present invention to human prostatecancer cell line LAPC-4 versus a control using only a solvent.

FIG. 21 depicts the dose response curves generated by exposing variousconcentrations of compound 7d of the present invention to human prostatecancer cell line LAPC-4 versus a control using only a solvent.

FIG. 22 depicts the dose response curves generated by exposing variousconcentrations of compound 8h of the present invention to human prostatecancer cell line LAPC-4 versus a control using only a solvent.

FIG. 23 depicts the dose response curves generated by exposing variousconcentrations of compound 1 of the present invention to human prostatecancer cell line CWR22R versus a control using only a solvent.

FIG. 24 depicts the dose response curves generated by exposing variousconcentrations of compound 6l of the present invention to human prostatecancer cell line CWR22R versus a control using only a solvent.

FIG. 25 depicts the dose response curves generated by exposing variousconcentrations of compound 6h of the present invention to human prostatecancer cell line CWR22R versus a control using only a solvent.

FIG. 26 depicts the dose response curves generated by exposing variousconcentrations of compound 6n of the present invention to human prostatecancer cell line CWR22R versus a control using only a solvent.

FIG. 27 depicts the dose response curves generated by exposing variousconcentrations of compound 7d of the present invention to human prostatecancer cell line CWR22R versus a control using only a solvent.

FIG. 28 depicts the dose response curves generated by exposing variousconcentrations of compound 8h of the present invention to human prostatecancer cell line CWR22R versus a control using only a solvent.

FIG. 29 depicts the dose response curves generated by exposing variousconcentrations of compound 1 of the present invention to human prostatecancer cell line LnCaP versus a control using only a solvent.

FIG. 30 depicts the dose response curves generated by exposing variousconcentrations of compound 6l of the present invention to human prostatecancer cell line LnCaP versus a control using only a solvent.

FIG. 31 depicts the dose response curves generated by exposing variousconcentrations of compound 6h of the present invention to human prostatecancer cell line LnCaP versus a control using only a solvent.

FIG. 32 depicts the dose response curves generated by exposing variousconcentrations of compound 6n of the present invention to human prostatecancer cell line LnCaP versus a control using only a solvent.

FIG. 33 depicts the dose response curves generated by exposing variousconcentrations of compound 7d of the present invention to human prostatecancer cell line LnCaP versus a control using only a solvent.

FIG. 34 depicts the dose response curves generated by exposing variousconcentrations of compound 8h of the present invention to human prostatecancer cell line LnCaP versus a control using only a solvent.

DETAILED DESCRIPTION OF THE INVENTION Abbreviations and Definitions

The abbreviations used herein have their conventional meaning within thechemical and biological arts.

Where moieties are specified by their conventional chemical formulae,written from left to right, they equally encompass the chemicallyidentical moieties that would result from writing the structure fromright to left, e.g., —CH₂O— is equivalent to —OCH₂—.

The term “alkyl,” by itself or as part of another substituent, means,unless otherwise stated, a straight (i.e. unbranched) or branched chain,or cyclic hydrocarbon radical, or combination thereof, which may befully saturated, mono- or polyunsaturated and can include di- andmultivalent radicals, having the number of carbon atoms designated (i.e.C₁-C₁₀ means one to ten carbons). Examples of saturated hydrocarbonradicals include, but are not limited to, groups such as methyl, ethyl,n-propyl, isopropyl, n-butyl, t-butyl, isobutyl, sec-butyl, cyclohexyl,(cyclohexyl)methyl, cyclopropylmethyl, homologs and isomers of, forexample, n-pentyl, n-hexyl, n-heptyl, n-octyl, and the like. Anunsaturated alkyl group is one having one or more double bonds or triplebonds. Examples of unsaturated alkyl groups include, but are not limitedto, vinyl, 2-propenyl, crotyl, 2-isopentenyl, 2-(butadienyl),2,4-pentadienyl, 3-(1,4-pentadienyl), ethynyl, 1- and 3-propynyl,3-butynyl, and the higher homologs and isomers.

The term “alkylene” by itself or as part of another substituent means adivalent radical derived from an alkyl, as exemplified, but not limited,by —CH₂CH₂CH₂CH₂—. Typically, an alkyl (or alkylene) group will havefrom 1 to 24 carbon atoms, including those groups having 10 or fewercarbon atoms. A “lower alkyl” or “lower alkylene” is a shorter chainalkyl or alkylene group, generally having eight or fewer carbon atoms.

The terms “alkoxy,” “alkylamino,” and “alkylthio” (or thioalkoxy) areused in their conventional sense, and refer to those alkyl groupsattached to the remainder of the molecule via an oxygen atom, an aminogroup, or a sulfur atom, respectively.

The term “heteroalkyl,” by itself or in combination with another term,means, unless otherwise stated, a stable straight or branched chain, orcyclic hydrocarbon radical, or combinations thereof, consisting of thestated number of carbon atoms and a heteroatom selected from the groupconsisting of O, N, P, Si and S, and wherein the nitrogen and sulfuratoms may optionally be oxidized and the nitrogen heteroatom mayoptionally be quaternized. The heteroatom(s) O, N, P and S and Si may beplaced at any interior position of the heteroalkyl group or at theposition at which the alkyl group is attached to the remainder of themolecule. Examples include, but are not limited to, —CH₂—CH₂—O—CH₃,—CH₂—CH₂—NH—CH₃, —CH₂—CH₂—N(CH₃)—CH₃, —CH₂—S—CH₂—CH₃, —CH₂—CH₂,—S(O)—CH₃, —CH₂—CH₂—S(O)₂—CH₃, —CH═CH—O—CH₃, —Si(CH₃)₃,—CH═CH—N(CH₃)—CH₃, O—CH₃, —O—CH₂—CH₃, and —CN. Up to two heteroatoms maybe consecutive, such as, for example, —CH₂—NH—OCH₃ and —CH₂—O—Si(CH₃)₃.Similarly, the term “heteroalkylene” by itself or as part of anothersubstituent means a divalent radical derived from heteroalkyl, asexemplified, but not limited by, —CH₂—CH₂—S—CH₂—CH₂— and—CH₂—S—CH₂—CH₂—NH—CH₂—. For heteroalkylene groups, heteroatoms can alsooccupy either or both of the chain termini (e.g., alkyleneoxy,alkylenedioxy, alkyleneamino, alkylenediamino, and the like). Stillfurther, for alkylene and heteroalkylene linking groups, no orientationof the linking group is implied by the direction in which the formula ofthe linking group is written. For example, the formula —C(O)₂R′—represents both —C(O)₂R′— and —R′C(O)₂—. As described above, heteroalkylgroups, as used herein, include those groups that are attached to theremainder of the molecule through a heteroatom, such as —C(O)R′,—C(O)NR′, —NR′R″, —OR′, —SR′, and/or —SO₂R′. Where “heteroalkyl” isrecited, followed by recitations of specific heteroalkyl groups, such as—NR′R″ or the like, it will be understood that the terms heteroalkyl and—NR′R″ are not redundant or mutually exclusive. Rather, the specificheteroalkyl groups are recited to add clarity. Thus, the term“heteroalkyl” should not be interpreted herein as excluding specificheteroalkyl groups, such as —NR′R″ or the like.

The terms “cycloalkyl” and “heterocycloalkyl”, by themselves or incombination with other terms, represent, unless otherwise stated, cyclicversions of “alkyl” and “heteroalkyl”, respectively. Additionally, forheterocycloalkyl, a heteroatom can occupy the position at which theheterocycle is attached to the remainder of the molecule. Examples ofcycloalkyl include, but are not limited to, cyclopentyl, cyclohexyl,1-cyclohexenyl, 3-cyclohexenyl, cycloheptyl, and the like. Examples ofheterocycloalkyl include, but are not limited to,1-(1,2,5,6-tetrahydropyridyl), 1-piperidinyl, 2-piperidinyl,3-piperidinyl, 4-morpholinyl, 3-morpholinyl, tetrahydrofuran-2-yl,tetrahydrofuran-3-yl, tetrahydrothien-2-yl, tetrahydrothien-3-yl,1-piperazinyl, 2-piperazinyl, and the like.

The terms “halo” or “halogen,” by themselves or as part of anothersubstituent, mean, unless otherwise stated, a fluorine, chlorine,bromine, or iodine atom. Additionally, terms such as “haloalkyl,” aremeant to include monohaloalkyl and polyhaloalkyl. For example, the term“halo(C₁-C₄)alkyl” is mean to include, but not be limited to,trifluoromethyl, 2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, andthe like.

The term “aryl” means, unless otherwise stated, a polyunsaturated,aromatic, hydrocarbon substituent which can be a single ring or multiplerings (preferably from 1 to 3 rings) which are fused together or linkedcovalently. The term “heteroaryl” refers to aryl groups (or rings) thatcontain from one to four heteroatoms selected from N, O, and S, whereinthe nitrogen and sulfur atoms are optionally oxidized, and the nitrogenatom(s) are optionally quaternized. A heteroaryl group can be attachedto the remainder of the molecule through a carbon or heteroatom.Non-limiting examples of aryl and heteroaryl groups include phenyl,1-naphthyl, 2-naphthyl, 4-biphenyl, 1-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl,3-pyrazolyl, 2-imidazolyl, 4-imidazolyl, pyrazinyl, 2-oxazolyl,4-oxazolyl, 2-phenyl-4-oxazolyl, 5-oxazolyl, 3-isoxazolyl, 4-isoxazolyl,5-isoxazolyl, 2-thiazolyl, 4-thiazolyl, 5-thiazolyl, 2-furyl, 3-furyl,2-thienyl, 3-thienyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-pyrimidyl,4-pyrimidyl, 5-benzothiazolyl, purinyl, 2-benzimidazolyl, 5-indolyl,1-isoquinolyl, 5-isoquinolyl, 2-quinoxalinyl, 5-quinoxalinyl,3-quinolyl, and 6-quinolyl. Substituent moieties for each of the abovenoted aryl and heteroaryl ring systems may be selected from the group ofacceptable substituent moieties described below.

For brevity, the term “aryl” when used in combination with other terms(e.g., aryloxy, arylthioxy, arylalkyl) includes both aryl and heteroarylrings as defined above. Thus, the term “arylalkyl” is meant to includethose radicals in which an aryl group is attached to an alkyl group(e.g., benzyl, phenethyl, pyridylmethyl and the like) including thosealkyl groups in which a carbon atom (e.g., a methylene group) has beenreplaced by, for example, an oxygen atom (e.g., phenoxymethyl,2-pyridyloxymethyl, 3-(1-naphthyloxy)propyl, and the like).

The term “oxo” as used herein means an oxygen that is double bonded to acarbon atom.

Each of the above terms (e.g., “alkyl,” “heteroalkyl,” “aryl” and“heteroaryl”) are meant to include both substituted and unsubstitutedforms of the indicated radical. Preferred substituent moieties for eachtype of radical are provided below.

Substituent moieties for the alkyl and heteroalkyl radicals (includingthose groups often referred to as alkylene, alkenyl, heteroalkylene,heteroalkenyl, alkynyl, cycloalkyl, heterocycloalkyl, cycloalkenyl, andheterocycloalkenyl) can be one or more of a variety of groups selectedfrom, but not limited to: —OR′, ═O, ═NR′, ═N—OR′, —NR′R″, —SR′,-halogen, —SiR′R″R′″, —OC(O)R′, —C(O)R′, —CO₂R′, —CONR′R″, —OC(O)NR′R″,—NR″C(O)R′, —NR′—C(O)NR″R′″, —NR″C(O)₂R′, —NR—C(NR′R″R′″)═NR″,—NR—C(NR′R″)═NR′″, —S(O)R′, —S(O)₂R′, —S(O)₂NR′R″, —NRSO₂R′, —CN and—NO₂ in a number ranging from zero to (2m′+1), where m′ is the totalnumber of carbon atoms in such radical. R′, R″, R′″ and R″″ eachpreferably independently refer to hydrogen, substituted or unsubstitutedheteroalkyl, substituted or unsubstituted cycloalkyl, substituted orunsubstituted heterocycloalkyl, substituted or unsubstituted aryl (e.g.,aryl substituted with 1-3 halogens), substituted or unsubstituted alkyl,alkoxy or thioalkoxy groups, or arylalkyl groups. When a compound of theinvention includes more than one R group, for example, each of the Rgroups is independently selected as are each R′, R″, R′″ and R″″ groupswhen more than one of these groups is present. When R′ and R″ areattached to the same nitrogen atom, they can be combined with thenitrogen atom to form a 4-, 5-, 6-, or 7-membered ring. For example,—NR′R″ is meant to include, but not be limited to, 1-pyrrolidinyl and4-morpholinyl. From the above discussion of substituent moieties, one ofskill, in the art will understand that the term “alkyl” is meant toinclude groups including carbon atoms bound to groups other thanhydrogen groups, such as haloalkyl (e.g., —CF₃ and —CH₂CF₃) and acyl(e.g., —C(O)CH₃, —C(O)CF₃, —C(O)CH₂OCH₃, and the like).

Similar to the substituent moieties described for the alkyl radical,substituent moieties for the aryl and heteroaryl groups are varied andmay be selected from, for example: halogen, —OR′, —NR′R″, —SR′,-halogen, —SiR′R″R′″, —OC(O)R′, —C(O)R′, —CONR′R″, —OC(O)NR′R″,—NR″C(O)R′, —NR″C(O)NR″R″, —NR″C(O)₂R′, —NR—C(NR′R″R″)═NR″″,—NR—C(NR′R″)═NR′″, —S(O)R′, —S(O)₂R′, —S(O)₂NR′R″, —NRSO₂R′, —CN and—NO₂, —R′, —N₃, —CH(Ph)₂, fluoro(C₁-C₄)alkoxy, and fluoro(C₁-C₄)alkyl,in a number ranging from zero to the total number of open valences onthe aromatic ring system; and where R′, R″, R″′ and R″″ are preferablyindependently selected from hydrogen, substituted or unsubstitutedalkyl, substituted or unsubstituted heteroalkyl, substituted orunsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl,substituted or unsubstituted aryl and substituted or unsubstitutedheteroaryl. When a compound of the invention includes more than one Rgroup, for example, each of the R groups is independently selected asare each R′, R″, R′″ and R″″ groups when more than one of these groupsis present.

Two of the substituent moieties on adjacent atoms of the aryl orheteroaryl ring may optionally form a ring of the formula-T-C(O)—(CRR′)_(q)-U-, wherein T and U are independently —NR—, —O—,—CRR′— or a single bond, and q is an integer of from 0 to 3.Alternatively, two of the substituent moieties on adjacent atoms of thearyl or heteroaryl ring may optionally be replaced with a substituent ofthe formula -A-(CH₂)_(r)—B—, wherein A and B are independently —CRR′—,—O—, —NR—, —S—, —S(O)—, —S(O)₂—, —S(O)₂NR′— or a single bond, and r isan integer of from 1 to 4. One of the single bonds of the new ring soformed may optionally be replaced with a double bond. Alternatively, twoof the substituent moieties on adjacent atoms of the aryl or heteroarylring may optionally be replaced with a substituent of the formula—(CRR′)_(s)—X′—(C′″R′″)_(d)—, where s and d are independently integersof from 0 to 3, and X′ is —O—, —NR′—, —S—, —S(O)—, —S(O)₂—, or—S(O)₂NR′—. The substituent moieties R, R′, R″ and R′″ are preferablyindependently selected from hydrogen, substituted or unsubstitutedalkyl, substituted or unsubstituted cycloalkyl, substituted orunsubstituted heterocycloalkyl, substituted or unsubstituted aryl, andsubstituted or unsubstituted heteroaryl.

As used herein, the term “heteroatom” or “ring heteroatom” is meant toinclude oxygen (O), nitrogen (N), sulfur (S), phosphorus (P), andsilicon (Si).

The term “pharmaceutically acceptable salts” is meant to include saltsof the active compounds which are prepared with relatively nontoxicacids or bases, depending on the particular substituent moieties foundon the compounds described herein. When compounds of the presentinvention contain relatively acidic functionalities, base addition saltscan be obtained by contacting the neutral form of such compounds with asufficient amount of the desired base, either neat or in a suitableinert solvent. Examples of pharmaceutically acceptable base additionsalts include Sodium, potassium, calcium, ammonium, organic amino, ormagnesium salt, or a similar salt. When compounds of the presentinvention contain relatively basic functionalities, acid addition saltscan be obtained by contacting the neutral form of such compounds with asufficient amount of the desired acid, either neat or in a suitableinert solvent. Examples of pharmaceutically acceptable acid additionsalts include those derived from inorganic acids like hydrochloric,hydrobromic, nitric, carbonic, monohydrogencarbonic, phosphoric,monohydrogenphosphoric, dihydrogenphosphoric, sulfuric,monohydrogensulfuric, hydriodic, or phosphorous acids and the like, aswell as the salts derived from relatively nontoxic organic acids likeacetic, propionic, isobutyric, maleic, malonic, benzoic, succinic,suberic, fumaric, lactic, mandelic, phthalic, benzenesulfonic,p-tolylsulfonic, citric, tartaric, methanesulfonic, and the like. Alsoincluded are salts of amino acids such as arginate and the like, andsalts of organic acids like glucuronic or galactunoric acids and thelike (see, for example, Berge et al., “Pharmaceutical Salts”, Journal ofPharmaceutical Science, 1977, 66, 1-19). Certain specific compounds ofthe present invention contain both basic and acidic functionalities thatallow the compounds to be converted into either base or acid additionsalts.

The neutral forms of the compounds are preferably regenerated bycontacting the salt with a base or acid and isolating the parentcompound in the conventional manner. The parent form of the compounddiffers from the various salt forms in certain physical properties, suchas solubility in polar solvents.

In addition to salt forms, the present invention provides compounds,which are in a prodrug form. Prodrugs of the compounds described hereinare those compounds that readily undergo chemical changes underphysiological conditions to provide the compounds of the presentinvention. Additionally, prodrugs can be converted to the compounds ofthe present invention by chemical or biochemical methods in an ex vivoenvironment. For example, prodrugs can be slowly converted to thecompounds of the present invention when placed in a transdermal patchreservoir with a suitable enzyme or chemical reagent.

Certain compounds of the present invention can exist in unsolvated formsas well as solvated forms, including hydrated forms. In general, thesolvated forms are equivalent to unsolvated forms and are encompassedwithin the scope of the present invention. Certain compounds of thepresent invention may exist in multiple crystalline or amorphous forms.In general, all physical forms are equivalent for the uses contemplatedby the present invention and are intended to be within the scope of thepresent invention.

Certain compounds of the present invention possess asymmetric carbonatoms (optical centers) or double bonds; the racemates, diastereomers,tautomers, geometric isomers and individual isomers are encompassedwithin the scope of the present invention. The compounds of the presentinvention do not include those which are known in the art to be toounstable to synthesize and/or isolate.

The compounds of the present invention may also contain unnaturalproportions of atomic isotopes at one or more of the atoms thatconstitute such compounds. For example, the compounds may beradiolabeled with radioactive isotopes, such as for example tritium(³H), iodine-125 (¹²⁵I) or carbon-14 (¹⁴C). All isotopic variations ofthe compounds of the present invention, whether radioactive or not, areencompassed within the scope of the present invention.

Where two groups are “optionally joined together to form a ring,” thetwo groups are covalently bonded together with the atom or atoms towhich the two groups are joined to form a substituted or unsubstitutedaryl, a substituted or unsubstituted heteroaryl, a substituted orunsubstituted cycloalkyl, or a substituted or unsubstitutedheterocycloalkyl ring.

The terms “arylalkyl,” “heteroarylalkyl,” “cycloalkyl-alkyl,” and“heterocycloalkyl-alkyl,” as used herein, refer to an aryl, heteroaryl,cycloalkyl and heterocycloalkyl, respectively, attached to the remainderof the molecule via an alkylene group. Where an “arylalkyl,”“heteroarylalkyl,” “cycloalkyl-alkyl,” or “heterocycloalkyl-alkyl” issubstituted, one or more substituent moieties may be covalently bondedto the alkylene moiety and/or the aryl, heteroaryl, cycloalkyl andheterocycloalkyl moieties, respectively. A “C₁-C₂₀” arylalkyl,heteroarylalkyl, cycloalkyl-alkyl, or heterocycloalkyl-alkyl, aremoieties in which a C₁-C₂₀ alkylene links an aryl, heteroaryl, C₄-C₈cycloalkyl, and 4 to 8 membered heterocycloalkyl, respectively, to theremainder of the molecule. A “C₁-C₈” arylalkyl, heteroarylalkyl,cycloalkyl-alkyl, or heterocycloalkyl-alkyl, are moieties in which aC₁-C₈ alkylene links an aryl, heteroaryl, C₅-C₇ cycloalkyl, and 5 to 7membered heterocycloalkyl, respectively, to the remainder of themolecule

A “substituent group,” as used herein, means a group selected from thefollowing moieties:

-   -   (A) —OH, —NH₂, —SH, —CN, —CF₃, oxy, halogen, unsubstituted        alkyl, unsubstituted heteroalkyl, unsubstituted cycloalkyl,        unsubstituted heterocycloalkyl, unsubstituted aryl,        unsubstituted heteroaryl, and    -   (B) alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, and        heteroaryl, substituted with at least one substituent selected        from:        -   (i) oxy, —OH, —NH₂, —SH, —CN, —CF₃, halogen, unsubstituted            alkyl, unsubstituted heteroalkyl, unsubstituted cycloalkyl,            unsubstituted heterocycloalkyl, unsubstituted aryl,            unsubstituted heteroaryl, and        -   (ii) alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl,            and heteroaryl, substituted with at least one substituent            selected from:            -   (a) oxy, —OH, —NH₂, —SH, —CN, —CF₃, halogen,                unsubstituted alkyl, unsubstituted heteroalkyl,                unsubstituted cycloalkyl, unsubstituted                heterocycloalkyl, unsubstituted aryl, unsubstituted                heteroaryl, and            -   (b) alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl,                aryl, or heteroaryl, substituted with at least one                substituent selected from oxy, —OH, —NH₂, —SH, —CN,                —CF₃, halogen, unsubstituted alkyl, unsubstituted                heteroalkyl, unsubstituted cycloalkyl, unsubstituted                heterocycloalkyl, unsubstituted aryl, and unsubstituted                heteroaryl.

A “size-limited substituent” or “size-limited substituent group,” asused herein means a group selected from all of the substituentsdescribed above for a “substituent group,” wherein each substituted orunsubstituted alkyl is a substituted or unsubstituted C₁-C₂₀ alkyl, eachsubstituted or unsubstituted heteroalkyl is a substituted orunsubstituted 2 to 20 membered heteroalkyl, each substituted orunsubstituted cycloalkyl is a substituted or unsubstituted C₄-C₈cycloalkyl, and each substituted or unsubstituted heterocycloalkyl is asubstituted or unsubstituted 4 to 8 membered heterocycloalkyl.

A “lower substituent” or “lower substituent group,” as used herein meansa group selected from all of the substituents described above for a“substituent group,” wherein each substituted or unsubstituted alkyl isa substituted or unsubstituted C₁-C₈ alkyl, each substituted orunsubstituted heteroalkyl is a substituted or unsubstituted 2 to 8membered heteroalkyl, each substituted or unsubstituted cycloalkyl is asubstituted or unsubstituted C₅-C₇ cycloalkyl, and each substituted orunsubstituted heterocycloalkyl is a substituted or unsubstituted 5 to 7membered heterocycloalkyl.

The term “treating” refers to any indicia of success in the treatment oramelioration of an injury, pathology or condition, including anyobjective or subjective parameter such as abatement; remission;diminishing of symptoms or making the injury, pathology or conditionmore tolerable to the patient; slowing in the rate of degeneration ordecline; making the final point of degeneration less debilitating;improving a patient's physical or mental well-being. The treatment oramelioration of symptoms can be based on objective or subjectiveparameters; including the results of a physical examination,neuropsychiatric exams, and/or a psychiatric evaluation. For example,the methods of the invention successfully treat a patient's delirium bydecreasing the incidence of disturbances in consciousness or cognition.

The term “higher alkyl” refers to those alkyl groups having at least sixcarbon atoms. The term “lower alkyl” refers to those alkyl groups havingfrom one to five carbon atoms.

DESCRIPTION OF THE EMBODIMENTS I. Compounds of the Present Invention

In one aspect, the present invention provides an antiproliferative,water soluble compound with high bioavailability having the generalformula (I)

Where A, B, and C are independently substituted alkyl, cycloalkyl, aryl,heterocycle, or substituted phenyl fused with a saturated or unsaturated4- to 6-membered rings optionally containing one to three heteroatomsindependently selected from N, O, and S; ‘X’ is oxygen or sulfur; ‘Y’ issulfur, sulfoxide or sulfone; X₁ is CO or hydrogen; R₅ is hydrogen, oralkyl or aryl group; X₂ is hydrogen or SR₆, wherein R₆ is a substitutedalkyl, cycloalkyl, aryl, heterocycle, or substituted phenyl fused with asaturated or unsaturated 4- to 6-membered rings optionally containingone to three heteroatoms independently selected from N, O, and S.

The present invention is further defined by compounds having the generalformulas (II)-(XIX):

Where R₁, R₃ is F, Cl, Br, I, CN, NO₂, NH₂, NHMe, NMe₂, CF₃, COOH,CONHOH, alkyl or aryl, group; R₅ is alkyl or aryl group; R₄ issubstituted pyridine, pyrimidine, thiazole, pyrrazole, benzothiozole,indole, benzofuran or any heterocyclic ring; ‘X’ is oxygen or sulfur;‘Y’ is sulfur, sulfoxide or sulfone; n is any integer.

Where R₁, R₂ is H, F, Cl, Br, I, CN, NO₂, NH₂, NHMe, NMe2, CF₃, COOH,CONHOH, alkyl or aryl, group; R₃ is substituted pyridine, pyrimidine,thiazole, pyrazole, benzothiozole, indole, benzofuran or anyheterocyclic ring; ‘X’ is oxygen or sulfur; ‘Y’ is sulfur, sulfoxide orsulfone; n is any integer.

Where A, B, and D are independently substituted alkyl, cycloalkyl, aryl,heterocycle, or substituted phenyl fused with a saturated or unsaturated4- to 6-membered rings optionally containing one to three heteroatomsindependently selected from N, O, and S; ‘X’ is oxygen or sulfur; ‘Y’ issulfur, sulfoxide or sulfone.

Where R₁, R₂ is H, F, Cl, Br, I, CN, NO₂, NH₂, NHMe, NMe₂, CF₃, COON,CONHOH, alkyl or aryl, group; R₃ is substituted pyridine, pyrimidine,thiazole, pyrrazole, benzothiozole, indole, benzofuran or anyheterocyclic ring; R₄, R₅ is substituted alkyl or aryl group; ‘X’ isoxygen or sulfur.

Where A, B, D, E and F are independently substituted alkyl, cycloalkyl,aryl, heterocycle, or substituted phenyl fused with a saturated orunsaturated 4- to 6-membered rings optionally containing one to threeheteroatoms independently selected from N, O, and S; ‘X’ is oxygen orsulfur.

Where A, B, D, E and G are independently substituted alkyl, cycloalkyl,aryl, heterocycle, or substituted phenyl fused with a saturated orunsaturated 4- to 6-membered rings optionally containing one to threeheteroatoms independently selected from N, O, and S; ‘X’ is oxygen orsulfur.

In another embodiment, the compounds of the present invention have theformulas provided in Table 1. these compounds were prepared according tothe synthesis schemes that are described below.

TABLE 1 Com- pound Structure  6a

 6b

 6c

 6d

 6e

 6f

 6g

 6h

 6i

 6j

 6k

 6l

 6m

 6n

 7a

 7b

 7c

 7d

 8a

 8b

 8c

 8d

 8e

 8f

 8g

 8h

 9

10

General Method for Synthesis of BPU Sulfur Analogs

These new analogs were synthesized in excellent yield by couplingcorresponding benzoylisocyanate or benzoylisothiocyanate and anilinederivatives. Sulfur analogs of aniline intermediate were synthesized byreaction of substituted amino thiophenol with aryl halide. Sulfoxide andsulfone derivatives were prepared by oxidation of sulfides (FIG. 1). Allthe compounds were characterized by NMR and LC-MS studies.

Chemistry. Novel sulfur derivatives of BPU were synthesized as shown inFIG. 1. Aniline derivatives 4 were prepared by reaction of substitutedaminophenols or amino thiophenols with substituted 2-chloropyrimidinesin the presence of K₂CO₃ and DMSO.8 Condensation of substituted benzoylisocyanates or benzoyl isothiocyanates with aniline derivatives 4 gave aseries of BPU and benzoylphenylthiourea (BPTU) analogues 6a-n. Reductionof the 2-NO₂ group was performed using Fe and AcOH 14 to obtain the2-NH₂ derivatives 7a-d. Sulfoxide and sulfone derivatives 8a-h wereprepared from the corresponding sulfide by oxidation with MCPBA(Heynderickx, A., et al., Synthesis, 1112-1116 (2003)) followed bychromatographic purification. All the compounds were characterized byspectral data analysis that confirmed the assigned structures.

Ring-A aniline modified amino acid prodrugs can prepared according toFIG. 2. The synthetic strategy involves coupling of Boc-protected aminoacid with ring-A amine on the BPU compounds using coupling reagent1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (WSCI.HCl).Finally Boc protection can be removed by treating with TFA. We havecompleted the synthesis of N,N-dimethylglycyl derivative of 7d. Otheramino acid derivatives synthesis is in progress and their activity willbe compared.

Ring-A aniline modified carbamylosulfenyl derivative of BPU sulfuranalog can be prepared according to FIG. 3. To prepare urea and benzoylurea modified prodrugs, scheme 4 will be used. Compound 15 can beprepared by treating 14 with oxalyl chloride and then it can be coupledwith 17 and 19 in dioxane to obtain prodrugs of general structure 20 and21 respectively. Representative example has been shown in FIG. 5. Bothring-A and urea modified prodrugs can be prepared according to FIG. 6.

BPU compounds have very low solubility in water and limited solubilityin common organic solvents. Therefore, the modification of thesecompounds to improve their bioavailability is expected to lead tocompounds with better biological properties.

The lead prodrugs from the in vitro and in vivo studies will be thensubjected to pharmacokinetic analysis to study stability, clearance andmetabolism profiles.

II. Assays Cytotoxic Evaluation of BPU Sulfur Analogs

Eight of these analogs 6d, 6h, 6n, 6g, 7b, 7d, 8g, and 8h were testedfor their cytotoxicity in seven pancreatic cell lines ASPC1, Panel,Panc203, Panc430, Pane 1005, MiaPaca2, and HS766T using MTT assay. Cellswere trypsinized, seeded at 5×103 cells/well in 96-well plate andallowed to grow for 24 hours before the treatment with exponentialincreasing concentrations of drugs in the presence of 10% FBS. After a96-hours period of treatment, 20 μl of MTT solution (5 mg/ml in PBS)were added to each well, and the plates were then incubated for 3 hoursat 37° C. Medium was then replaced with 100 μl of DMSO per well. Plateswere shaken and the optical density was measured at 570 nm using amultiwell plate reader (Bio-Rad, Model 550, Bio-Rad Inc., Hercules,Calif.). Each experiment was performed in triplicate for each drugconcentration and was carried out independently at least 3 times. TheIC₅₀ value was defined as the concentration needed for a 50% reductionin the absorbance calculated based on the survival curves. Response todrug treatment was assessed by standardizing treatment groups tountreated controls. Compounds activity was compared with compound 1. Twocompounds 6n and 7d found to be 20 time more potent than compound 1.

Growth Inhibition Results

Growth inhibition of pancreatic cell lines by compound 1 and thecompounds 6n, 7d, 6d, 6h, 7b, 6g, 8g, 8h of the present invention aredepicted in following FIGS. 8-16, respectively.

Growth Inhibition Assay for Prostate Cancer Cell Lines

This MTT assay was carried using human prostate cancer cell linesCWR22R, LAPC-4 and LnCap using Promega cell titer 96 nonradioactive cellproliferation kit. Cells were trypsinized and centrifuged at 1000 rpmfor 5 minutes and the supernatant was aspirated. Cells were resuspendedwith HBSS to a concentration of 106/ml. Cell suspension was diluted inserum containing medium to plate 2000 cells/well/0.2 ml. Cell suspensionwas transferred to sterile solution basin. Cells were plated usingmultichannel pipette set at 200 μl and incubated overnight forattachment. One plate was prepared for day 0 MTT assay as follows: 100μl of media was removed from each well using multichannel pipette and 15μl of MTT reagent was added. Plates were incubated at 37° C. for 4 hrs.100 μl of STOP solution was added to each well and incubated at roomtemperature for 1 hour. Absorbance was measured at 570 and 650 nm onplate reader. All the media from the remaining plates were carefullyremoved and the cells were treated with the media containing drug. MTTassays were done on days 0, 3, 5 and 7. In the preliminary screeningfour BPU sulfur analogs 6l, 6h, 6n, 7d and 8b were screened and theiractivity was compared with compound 1. Two compounds 6n and 7d found tobe 20 times more potent compare to compound 1 compound. The data isshown in the following FIGS. 17-34.

Biology. Table 2 (Growth Inhibition of Pancreatic Cancer Cell Lines byBPU and BPTU Analogues (IC₅₀)^(a)) compares the cytotoxicity of a seriesof new derivatives against a panel of seven pancreatic cancer celllines, as determined by MTT assay. See, Mosmann, T., J. Immunol. Methods65:55-63 (1983).

TABLE 2 compd ASPC1 Panc1 Panc203 Panc430 Panc1005 Miapaca2 HS766 1 0.850.70 0.95 0.80 5.00 0.85 1.00 6g >10 >10 7.50 2.00 >10 5.00 5.50 6h10.00 9.50 >10 7.50 >10 10.00 8.50 6m 0.77 0.68 >10 10.00 >10 0.74 0.476n 1.00 0.085 0.70 0.09 10.00 0.086 0.088 7b 6.0 7.5 5.5 6.5 >10 7.0 8.07c 5.20 3.50 >10 >10 >10 0.92 0.27 7d 0.45 0.09 0.085 0.20 10.00 0.0950.70 8g 2.50 6.50 5.50 5.50 5.50 8.00 6.50 8h >10 >10 >10 >10 >10 6.10>10^(a) IC₅₀ values expressed in μm; Average of three independentexperiments.

Of all the tested compounds, compound 6n possessed the highest potency,with IC₅₀ values 0.085, 0.09, 0.086, and 0.088 μM against the Panc1,Panc430, Miapaca2, and HS766 cell lines, respectively. Results withcompound 7d demonstrated that substitution of NH₂ for the NO₂ group (6n)in the benzoyl ring did not diminish the cytotoxic activity against thePanc1, Panc203, and Miapaca2 cell lines. Compounds 6n and 7d were 7- to11-fold more potent than 1 against these cell lines. In the presence of6m, the growth of cell lines ASPC1, Panc1, Miapaca2, and HS766 wasinhibited, with IC₅₀ values of 0.77, 0.68, 0.74, and 0.47 μM,respectively.

Cell lines Miapaca2 and HS766 were susceptible to 7c, with IC₅₀ valuesof 0.92 and 0.27 μM, respectively. Compound 8g was effective ininhibiting the growth of all the cell lines, with IC₅₀ values rangingfrom 2.5 to 8.0 μM. For the remaining compounds, the IC₅₀ was >10 μM forall the cell lines. Compounds with significant inhibitory activity inpancreatic cancer cells were further evaluated against prostate cancercell lines in the MTT assay, (Mosmann, T., J. Immunol. Methods 65:55-63(1983)) and the results were compared to those for compound 1 (Table 3).Table 3 demonstrates the growth inhibition of prostate cancer cell linesby BPU and BPTU Analogues (IC₅₀)^(a)

TABLE 3 compd CWR22R LnCaP LAPC-4 1 1.00 0.50 0.25 6h >10 >10 6.00 6I5.00 0.50 0.60 6m 5.00 1.00 0.20 6n 0.05 0.05 0.05 7c 2.00 0.75 0.75 7d0.50 0.50 0.10 8h >10 0.50 4.00^(a) IC₅₀ values expressed in μm; Average of three independentexperiments. See, Mosmann, T., J. Immunol. Methods 65:55-63 (1983)

Compound 6n was the most highly potent of the tested compounds, with anIC₅₀ of 0.05 μM against all prostate cell lines. It was 5- to 20-foldmore potent than compound 1. The CWR22 and LnCap cell lines were equallysensitive (IC₅₀) 0.5 μM) to the growth inhibitory effects of 7d. 7e alsoinhibited cell growth in the prostate cancer cell lines LnCap and LAPC-4(IC₅₀) 0.75 μM).

6n treatment of MCF-7 cells for 48 hours produced rates of apoptosis of29.7% (10 nM), 51.2% (30 nM), and 53.6% (100 nM), respectively, ascompared to 16.1%, 19.5%, and 27.7% for 1 at the same concentration.Apoptosis of MCF-7 cells by 6n and 1 are demonstrated in FIG. 7 and 6nwas more potent than 1 in killing MCF-7 cells (Zhu, T., et al., CancerRes, 65:317-324 (2005)).

A mechanism-based tubulin assembly analysis (Hamel, E., Cell Biochem.Biophys., 38:1-22 (2003); and Verdier-Pinard, P., et al., Mol.Pharmacol, 53:62-76 (1998)) of four compounds from this series was alsoconducted. As shown in Table 4, 6n and 6m were effective in inhibitingtubulin assembly, with IC₅₀ values of 2.1 and 4.7 μM, respectively.These values were comparable to the IC₅₀ of 1.7 μM for the drugscombretastatin and podophyllotoxin, and 2.7 μM for colchicine, whichhave a similar mechanism of action. Our data indicate (Table 4) thatnone of our compounds bound to the colchicine binding site on themicrotubules.

TABLE 4 Inhibition of Tubulin Polymerization and Colchicine Bindingtubulin^(a) IC₅₀ ± SD colchicine binding^(b) compd (μM)± (% ± SD) 6m 4.7± 0.3 21 ± 5 6n 2.1 ± 0.5 17 ± 3 7c  36 ± 0.7 nd c 7d >40 ndcombretastatin A4 1.7 ± 0.2 94 ± 5 podophyllotoxin 1.7 ± 0.1 77 ± 8colchicines 2.7 ± 0.4 ^(a)Inhibition of tubulin polymerization. Tubulinwas at 10 μM (Hamel, E., Cell Biochem. Biophys., 38: 1-22 (2003)).^(b)Inhibition of [³H]colchicine binding. Tubulin was at 1 μM; both[³H]colchicines and inhibitor were at 5 μM (Verdier-Pinard, P., et al.,Mol. Pharmacol, 53: 62-76 (1998)). c No data.

From Tables 2 and 3, it is interesting to note that the NO₂ and NH₂substituents at position 2 of the benzoyl moiety and the Br substituentat position 4 on the pyrimidinyl ring apparently play an important rolein the activity of these compounds. Compounds lacking either or bothgroups displayed weak activity. These findings were in accordance withpreviously reported structure-activity relationships. See, Okada, H., etal., Chem. Pharm. Bull., 39:2308-2315 (1991); and Gurulingappa, H., etal., Bioorg. Med. Chem. Lett., 14:2213-2216 (2004). In addition,replacement of the urea moiety with thiourea (6g and 6h) had littleeffect on the activity. Introduction of a sulfide bridge between phenyland pyrimidinyl rings resulted in higher activity than did an etherlink. This variation was observed for compounds 6n and 7d (Tables 2 and3), both of which showed a significant anticancer effect. For example,an increase in potency of 111-fold for Panc430, 8-fold for Panel andMia-paca2, 14-fold for Panc203, 100-fold for CWR22R, and 20-fold for theLnCap cell line was obtained for compound 6n over 6m. Similarly, whencompared to 7c, the activity of 7d was increased by >117-fold forPanc203, 9-fold for Miapaca2, >50-fold for Panc430, 39-fold for Panel,and 7-fold for the LAPC-4 cell line. A large decrease in the activity of8g and 8h (when compared to that of 6n) was also seen with themodification of sulfide to sulfoxide and sulfone. Even though 6m and 6nwere effective in inhibiting tubulin assembly, their poor bindingaffinity for colchicine binding site indicates the existence of analternate binding site mechanism.

In conclusion, the sulfur analogues of BPU of the present invention hadexcellent growth inhibition activity against both pancreatic andprostate cancer cell lines. 6n and 7d were both found to be more potentthan compound 1.

In another aspect, the present invention provides pharmaceuticalcompositions. The pharmaceutical composition includes a pharmaceuticallyacceptable excipient and a compound having the formula (I-XXIII):

In one aspect, the present invention provides an antiproliferative,water soluble compound with high bioavailability having the generalformula (I)

Where A, B, and C are independently substituted alkyl, cycloalkyl, aryl,heterocycle, or substituted phenyl fused with a saturated or unsaturated4- to 6-membered rings optionally containing one to three heteroatomsindependently selected from N, O, and S; ‘X’ is oxygen or sulfur; ‘Y’ issulfur, sulfoxide or sulfone; X₁ is CO or hydrogen; R₅ is hydrogen, oralkyl or aryl group; X₂ is hydrogen or SR₆, wherein R₆ is a substitutedalkyl, cycloalkyl, aryl, heterocycle, or substituted phenyl fused with asaturated or unsaturated 4- to 6-membered rings optionally containingone to three heteroatoms independently selected from N, O, and S.

The present invention is further defined by compounds having the generalformulas (II)-(XIX):

Where R₁, R₃ is H, F, Cl, Br, I, CN, NO₂, NH₂, NHMe, NMc₂, CF₃, COOH,CONHOH, alkyl or aryl, group; R₅ is alkyl or aryl group; R₄ issubstituted pyridine, pyrimidine, thiazole, pyrrazole, benzothiozole,indole, benzofuran or any heterocyclic ring; ‘X’ is oxygen or sulfur;‘Y’ is sulfur, sulfoxide or sulfone; n is any integer.

Where R₁, R₂ is H, F, Cl, Br, I, CN, NO₂, NH₂, NHMe, NMe₂, CF₃, COOH,CONHOH, alkyl or aryl, group; R₃ is substituted pyridine, pyrimidine,thiazole, pyrrazole, benzothiozole, indole, benzofuran or anyheterocyclic ring; ‘X’ is oxygen or sulfur; ‘Y’ is sulfur, sulfoxide orsulfone; n is any integer.

Where A, B, and D are independently substituted alkyl, cycloalkyl, aryl,heterocycle, or substituted phenyl fused with a saturated or unsaturated4- to 6-membered rings optionally containing one to three heteroatomsindependently selected from N, O, and S; ‘X’ is oxygen or sulfur; ‘Y’ issulfur, sulfoxide or sulfone.

Where R₁, R₂ is H, F, Cl, Br, I, CN, NO₂, NH₂, NHMe, NMe₂, CF₃, COOH,CONHOH, alkyl or aryl, group; R₃ is substituted pyridine, pyrimidine,thiazole, pyrrazole, benzothiozole, indole, benzofuran or anyheterocyclic ring; R₄, R₅ is substituted alkyl or aryl group; ‘X’ isoxygen or sulfur.

Where A, B, D, E and F are independently substituted alkyl, cycloalkyl,aryl, heterocycle, or substituted phenyl fused with a saturated orunsaturated 4- to 6-membered rings optionally containing one to threeheteroatoms independently selected from N, O, and S; ‘X’ is oxygen orsulfur.

Where A, B, D, E and G are independently substituted alkyl, cycloalkyl,aryl, heterocycle, or substituted phenyl fused with a saturated orunsaturated 4- to 6-membered rings optionally containing one to threeheteroatoms independently selected from N, O, and S; ‘X’ is oxygen orsulfur.

The pharmaceutical compositions described herein are typically used forchemotherapy of various human illnesses, such as but not limited toosteoporesis, psoriasis, kidney failure, and immunosuppressant disordersin a subject in need of such treatment.

In an exemplary embodiment, the pharmaceutical composition includes from1 to 2000 milligrams of a compound of Formula (I-XXIII). In someembodiments, the pharmaceutical composition includes from 1 to 1500milligrams of the compound of Formula (I-XXIII). In other embodiments,the pharmaceutical composition includes from 1 to 1000 milligrams of thecompound of Formula (I-XXIII).

The compounds of the present invention can be prepared and administeredin a wide variety of oral, parenteral and topical dosage forms. Oralpreparations include tablets, pills, powder, dragees, capsules, liquids,lozenges, gels, syrups, slurries, suspensions, etc., suitable foringestion by the patient. The compounds of the present invention canalso be administered by injection, that is, intravenously,intramuscularly, intracutaneously, subcutaneously, intraduodenally, orintraperitoneally. Also, the compounds described herein can beadministered by inhalation, for example, intranasally. Additionally, thecompounds of the present invention can be administered transdermally.The compounds of this invention can also be administered by inintraocular, intravaginal, and intrarectal routes includingsuppositories, insufflation, powders and aerosol formulations (forexamples of steroid inhalants, see Rohatagi, J. Clin. Pharmacol.35:1187-1193, 1995; Tjwa, Ann. Allergy Asthma Immunol. 75:107-111,1995). Thus, the pharmaceutical compositions described herein may beadapted for oral administration. In some embodiments, the pharmaceuticalcomposition is in the form of a tablet. Moreover, the present inventionprovides pharmaceutical compositions including a pharmaceuticallyacceptable carrier or excipient and either a compound of Formula(I-XXIII), or a pharmaceutically acceptable salt of a compound ofFormula (I-XXIII).

For preparing pharmaceutical compositions from the compounds of thepresent invention, pharmaceutically acceptable carriers can be eithersolid or liquid. Solid form preparations include powders, tablets,pills, capsules, cachets, suppositories, and dispersible granules. Asolid carrier can be one or more substances, which may also act asdiluents, flavoring agents, binders, preservatives, tabletdisintegrating agents, or an encapsulating material. Details ontechniques for formulation and administration are well described in thescientific and patent literature, see, e.g., the latest edition ofRemington's Pharmaceutical Sciences, Maack Publishing Co, Easton Pa.(“Remington's”).

In powders, the carrier is a finely divided solid, which is in a mixturewith the finely divided active component. In tablets, the activecomponent is mixed with the carrier having the necessary bindingproperties in suitable proportions and compacted in the shape and sizedesired.

The powders and tablets preferably contain from 5% or 10% to 70% of theactive compound. Suitable carriers are magnesium carbonate, magnesiumstearate, talc, sugar, lactose, pectin, dextrin, starch, gelatin,tragacanth, methylcellulose, sodium carboxymethylcellulose, a lowmelting wax, cocoa butter, and the like. The term “preparation” isintended to include the formulation of the active compound withencapsulating material as a carrier providing a capsule in which theactive component with or without other carriers, is surrounded by acarrier, which is thus in association with it. Similarly, cachets andlozenges are included. Tablets, powders, capsules, pills, cachets, andlozenges can be used as solid dosage forms suitable for oraladministration.

Suitable solid excipients are carbohydrate or protein fillers include,but are not limited to sugars, including lactose, sucrose, mannitol, orsorbitol; starch from corn, wheat, rice, potato, or other plants;cellulose such as methyl cellulose, hydroxypropylmethyl-cellulose, orsodium carboxymethylcellulose; and gums including arabic and tragacanth;as well as proteins such as gelatin and collagen. If desired,disintegrating or solubilizing agents may be added, such as thecross-linked polyvinyl pyrrolidone, agar, alginic acid, or a saltthereof, such as sodium alginate.

Dragee cores are provided with suitable coatings such as concentratedsugar solutions, which may also contain gum arabic, talc,polyvinylpyrrolidone, carbopol gel, polyethylene glycol, and/or titaniumdioxide, lacquer solutions, and suitable organic solvents or solventmixtures. Dyestuffs or pigments may be added to the tablets or drageeliquid forms include solutions, suspensions, and emulsions. Thesepreparations may contain, in addition to the active component,colorants, flavors, stabilizers, buffers, artificial and naturalsweeteners, dispersants, thickeners, solubilizing agents, and the like.

Oil suspensions can be formulated by suspending a compound of thisinvention in a vegetable oil, such as arachis oil, olive oil, sesame oilor coconut oil, or in a mineral oil such as liquid paraffin; or amixture of these. The oil suspensions can contain a thickening agent,such as beeswax, hard paraffin or cetyl alcohol. Sweetening agents canbe added to provide a palatable oral preparation, such as glycerol,sorbitol or sucrose. These formulations can be preserved by the additionof an antioxidant such as ascorbic acid. As an example of an injectableoil vehicle, see Minto, J. Pharmacol. Exp. Ther., 281:93-102 (1997). Thepharmaceutical formulations of the invention can also be in the form ofoil-in-water emulsions. The oily phase can be a vegetable oil or amineral oil, described above, or a mixture of these. Suitableemulsifying agents include naturally-occurring gums, such as gum acaciaand gum tragacanth, naturally occurring phosphatides, such as soybeanlecithin, esters or partial esters derived from fatty acids and hexitolanhydrides, such as sorbitan mono-oleate, and condensation products ofthese partial esters with ethylene oxide, such as polyoxyethylenesorbitan mono-oleate. The emulsion can also contain sweetening agentsand flavoring agents, as in the formulation of syrups and elixirs. Suchformulations can also contain a demulcent, a preservative, or a coloringagent.

The compounds of this invention of the invention can be delivered bytransdermally, by a topical route, formulated as applicator sticks,solutions, suspensions, emulsions, gels, creams, ointments, pastes,jellies, paints, powders, and aerosols.

The compounds of this invention of the invention can also be deliveredas microspheres for slow release in the body. For example, microspherescan be administered via intradermal injection of drug-containingmicrospheres, which slowly release subcutaneously (see Rao, J. BiomaterSci. Polym. Ed., 7:623-645 (1995); as biodegradable and injectable gelformulations (see, e.g., Gao Pharm. Res., 12:857-863 (1995); or, asmicrospheres for oral administration (see, e.g., Eyles, J. Pharm.Pharmacol., 49:669-674 (1997). Both transdermal and intradermal routesafford constant delivery for weeks or months.

The compounds of this invention pharmaceutical formulations of theinvention can be provided as a salt and can be formed with many acids,including but not limited to hydrochloric, sulfuric, acetic, lactic,tartaric, malic, succinic, etc. Salts tend to be more soluble in aqueousor other protonic solvents that are the corresponding free base coatingsfor product identification or to characterize the quantity of activecompound (i.e., dosage). Pharmaceutical preparations of the inventioncan also be used orally using, for example, push-fit capsules made ofgelatin, as well as soft, sealed capsules made of gelatin and a coatingsuch as glycerol or sorbitol. Push-fit capsules can contain thecompounds of this invention mixed with a filler or binders such aslactose or starches, lubricants such as talc or magnesium stearate, and,optionally, stabilizers. In soft capsules, the compounds of thisinvention compounds may be dissolved or suspended in suitable liquids,such as fatty oils, liquid paraffin, or liquid polyethylene glycol withor without stabilizers.

For preparing suppositories, a low melting wax, such as a mixture offatty acid glycerides or cocoa butter, is first melted and the activecomponent is dispersed homogeneously therein, as by stirring. The moltenhomogeneous mixture is then poured into convenient sized molds, allowedto cool, and thereby to solidify.

Liquid form preparations include solutions, suspensions, and emulsions,for example, water or water/propylene glycol solutions. For parenteralinjection, liquid preparations can be formulated in solution in aqueouspolyethylene glycol solution.

Aqueous solutions suitable for oral use can be prepared by dissolvingthe active component in water and adding suitable colorants, flavors,stabilizers, and thickening agents as desired. Aqueous suspensionssuitable for oral use can be made by dispersing the finely dividedactive component in water with viscous material, such as natural orsynthetic gums, resins, methylcellulose, sodium carboxymethylcellulose,hydroxypropylmethylcellulose, sodium alginate, polyvinylpyrrolidone, gumtragacanth and gum acacia, and dispersing or wetting agents such as anaturally occurring phosphatide (e.g., lecithin), a condensation productof an alkylene oxide with a fatty acid (e.g., polyoxyethylene stearate),a condensation product of ethylene oxide with a long chain aliphaticalcohol (e.g., heptadecaethylene oxycetanol), a condensation product ofethylene oxide with a partial ester derived from a fatty acid and ahexitol (e.g., polyoxyethylene sorbitol mono-oleate), or a condensationproduct of ethylene oxide with a partial ester derived from fatty acidand a hexitol anhydride (e.g., polyoxyethylene sorbitan mono-oleate).The aqueous suspension can also contain one or more preservatives suchas ethyl or n-propyl p-hydroxybenzoate, one or more coloring agents, oneor more flavoring agents and one or more sweetening agents, such assucrose, aspartame or saccharin. Formulations can be adjusted forosmolarity.

Also included are solid form preparations, which are intended to beconverted, shortly before use, to liquid form preparations for oraladministration. Such forms. In other cases, the preparation may be alyophilized powder in 1 mM-50 mM histidine, 0.1%-2% sucrose, 2%-7%mannitol at a pH range of 4.5 to 5.5, that is combined with buffer priorto use.

In another embodiment, the compounds of this invention having theformulations of the invention are useful for parenteral administration,such as intravenous (IV) administration or administration into a bodycavity or lumen of an organ. The formulations for administration willcommonly comprise a solution of the compounds of this inventiondissolved in a pharmaceutically acceptable carrier. Among the acceptablevehicles and solvents that can be employed are water and Ringer'ssolution, an isotonic sodium chloride. In addition, sterile fixed oilscan conventionally be employed as a solvent or suspending medium. Forthis purpose any bland fixed oil can be employed including syntheticmono- or diglycerides. In addition, fatty acids such as oleic acid canlikewise be used in the preparation of injectables. These solutions aresterile and generally free of undesirable matter. These formulations maybe sterilized by conventional, well known sterilization techniques. Theformulations may contain pharmaceutically acceptable auxiliarysubstances as required to approximate physiological conditions such aspH adjusting and buffering agents, toxicity adjusting agents, e.g.,sodium acetate, sodium chloride, potassium chloride, calcium chloride,sodium lactate and the like. The concentration of the compounds of thisinvention in these formulations can vary widely, and will be selectedprimarily based on fluid volumes, viscosities, body weight, and thelike, in accordance with the particular mode of administration selectedand the patient's needs. For IV administration, the formulation can be asterile injectable preparation, such as a sterile injectable aqueous oroleaginous suspension. This suspension can be formulated according tothe known art using those suitable dispersing or wetting agents andsuspending agents. The sterile injectable preparation can also be asterile injectable solution or suspension in a nontoxicparenterally-acceptable diluent or solvent, such as a solution of1,3-butanediol.

In another embodiment, formulations having the compounds of thisinvention can be delivered by the use of liposomes which fuse with thecellular membrane or are endocytosed, i.e., by employing ligandsattached to the liposome, or attached directly to the oligonucleotide,that bind to surface membrane protein receptors of the cell resulting inendocytosis. By using liposomes, particularly where the liposome surfacecarries ligands specific for target cells, or are otherwisepreferentially directed to a specific organ, one can focus the deliveryof the compounds of this invention into the target cells in vivo. (See,e.g., Al-Muhammed, J. Microencapsul., 13:293-306, 1996; Chonn, Curr.Opin. Biotechnol., 6:698-708 (1995); Ostro, Am. J. Hosp. Pharm.,46:1576-1587 (1989).

The pharmaceutical preparation is preferably in unit dosage form. Insuch form the preparation is subdivided into unit doses containingappropriate quantities of the active component. The unit dosage form canbe a packaged preparation, the package containing discrete quantities ofpreparation, such as packeted tablets, capsules, and powders in vials orampoules. Also, the unit dosage form can be a capsule, tablet, cachet,or lozenge itself, or it can be the appropriate number of any of thesein packaged form.

The quantity of active component in a unit dose preparation may bevaried or adjusted from 0.1 mg to 10000 mg, more typically 1.0 mg to1000 mg, most typically 10 mg to 500 mg, according to the particularapplication and the potency of the active component. The compositioncan, if desired, also contain other compatible therapeutic agents.

II. Methods for Treating Conditions in Need of Chemotherapy

In still another aspect, the present invention provides a method for thetreatment of a disorder or condition which requires chemotherapy. Inthis method, a subject in need of such treatment is administered aneffective amount of a compound having one of the formulae providedabove. The amount is effective in treating the illness that the patientis afflicted with.

A variety of disease sates are capable of being treated withchemotherapy. Exemplary disease states include, but are not limited tocancer, kidney failure, osteoporosis, psoriasis, kidney failure, andimmunosuppressant disorders. The methods of treatment includesadministering to a patient in need of such treatment, a therapeuticallyeffective amount of a compound according to Formula (I-XXIII), or apharmaceutically acceptable salt thereof.

Thus, in an exemplary embodiment, the present invention provides amethod of treating a disorder or condition through chemotherapy, themethod including administering to a subject in need of such treatment,an effective amount of a compound of the present invention, such as acompound of Formula (I-XXIII).

The amount of the compounds of this invention adequate to treat adisease is defined as a “therapeutically effective dose”. The dosageschedule and amounts effective for this use, i.e., the “dosing regimen,”will depend upon a variety of factors, including the stage of thedisease or condition, the severity of the disease or condition, thegeneral state of the patient's health, the patient's physical status,age and the like. In calculating the dosage regimen for a patient, themode of administration also is taken into consideration.

The dosage regimen also takes into consideration pharmacokineticsparameters well known in the art, i.e., the rate of absorption,bioavailability, metabolism, clearance, and the like (see, e.g.,Hidalgo-Aragones, J. Steroid Biochem. Mol. Biol., 58:611-617 (1996);Groning, Pharmazie, 51:337-341 (1996); Fotherby, Contraception, 54:59-69(1996); Johnson, J. Pharm. Sci., 84:1144-1146 (1995); Rohatagi,Pharmazie, 50:610-613 (1995); Brophy, Eur. J. Clin. Pharmacol.,24:103-108 (1983); the latest Remington's, supra). The state of the artallows the clinician to determine the dosage regimen for each individualpatient, and disease or condition treated.

Single or multiple administrations of formulations having the compoundsof this invention can be administered depending on the dosage andfrequency as required and tolerated by the patient. The formulationsshould provide a sufficient quantity of active agent to effectivelytreat the disease state. Thus, in one embodiment, the pharmaceuticalformulations for oral administration of the compounds of this inventionis in a daily amount of between about 0.5 to about 20 mg per kilogram ofbody weight per day. In an alternative embodiment, dosages are fromabout 1 mg to about 4 mg per kg of body weight per patient per day areused. Lower dosages can be used, particularly when the drug isadministered to an anatomically secluded site, such as the cerebralspinal fluid (CSF) space, in contrast to administration orally, into theblood stream, into a body cavity or into a lumen of an organ.Substantially higher dosages can be used in topical administration.Actual methods for preparing parenterally administrable formulationshaving the compounds of this invention will be known or apparent tothose skilled in the art and are described in more detail in suchpublications as Remington's, supra. See also Nieman, In “ReceptorMediated Antisteroid Action,” Agarwal, et al., eds., De Gruyter, N.Y.(1987).

After a pharmaceutical composition including a compound of the inventionhas been formulated in an acceptable carrier, it can be placed in anappropriate container and labeled for treatment of an indicatedcondition. For administration of the compounds of this invention, suchlabeling would include, e.g., instructions concerning the amount,frequency and method of administration. In one embodiment, the inventionprovides for a kit for the treatment of cancer or kidney failure in ahuman which includes the compounds of this invention and instructionalmaterial teaching the indications, dosage and schedule of administrationof the compounds of this invention.

III. Exemplary Syntheses

The compounds of the invention are synthesized by an appropriatecombination of generally well known synthetic methods. Techniques usefulin synthesizing the compounds of the invention are both readily apparentand accessible to those of skill in the relevant art. The discussionbelow is offered to illustrate certain of the diverse methods availablefor use in assembling the compounds of the invention. However, thediscussion is not intended to define the scope of reactions or reactionsequences that are useful in preparing the compounds of the presentinvention.

IV. Examples

As can be appreciated by the skilled artisan, methods of synthesizingthe compounds of the described herein will be evident to those ofordinary skill in the art. Additionally, the various synthetic steps maybe performed in an alternate sequence or order to give the desiredcompounds. Synthetic chemistry transformations and protecting groupmethodologies (protection and deprotection) useful in synthesizing thecompounds described herein are known in the art and include, forexample, those such as described in R. Larock, Comprehensive OrganicTransformations, VCH Publishers (1989); T. W. Greene and P. G. M. Wuts,Protective Groups in Organic Synthesis, 2d. Ed., John Wiley and Sons(1991); L. Fieser and M. Fieser, Fieser and Fieser's Reagents forOrganic Synthesis, John Wiley and Sons (1994); and L. Paquette, ed.,Encyclopedia of Reagents for Organic Synthesis, John Wiley and Sons(1995), and subsequent editions thereof.

The discussion below is offered to illustrate certain of the diversemethods available for use in assembling the compounds of the invention.However, the discussion is not intended to define the scope of reactionsor reaction sequences that are useful in preparing the compounds of thepresent invention.

Experimental Section:

Unless otherwise noted, reactions were run in flame-dried round-bottomedflasks under an atmosphere of ultra-high-purity (UHP) argon. Allreactive liquid reagents were transferred by syringe or cannula and wereadded into the flask through a rubber septum. Tetrahydrofuran wasfreshly distilled from sodium benzophenone ketyl immediately prior touse. All other solvents and reagents were used as received unlessotherwise stated. n-BuLi was obtained from commercial sources and wastitrated with N-pivaloyl-O-toluidine prior to use. Diethyl ether (ether)and tetrahydrofuran (THF) were distilled from sodium benzophenone ketylprior to use. Methylene chloride (CH₂Cl₂) was distilled from calciumhydride prior to use. All other compounds were purchased from AldrichChemical Co. and used without further purification. Analyticalthin-layer chromatography (TLC) was conducted with silica gel 60 F254plates (250 lm thickness; Merck). Column chromatography was performedusing short path silica gel (particle size <230 mesh), flash silica gel(particle size 400 230 mesh), or Florisil (200 mesh). Yields are notoptimized. Purity of products was judged to be >95% based on theirchromatographic homogeneity. High-performance liquid chromatography(HPLC) was carried out with a Rainin HPLX system equipped with two 25mL/min preparative pump heads using Rainin Dynamax 10 250 mm(semipreparative) columns packed with 60 A ° silica gel (8 lm poresize), either as bare silica or as C-18-bonded silica. Melting pointswere measured using a MeI-Temp metal-block apparatus and wereuncorrected. Nuclear magnetic resonance (NMR) spectra were obtainedeither, on a Varian XL-400 spectrometer, operating at 400 MHz for ¹H and100 MHz for ¹³C, or on a Varian XL-500 spectrometer, operating at 500MHz for ¹H and 125 MHz for ¹³C. Chemical shifts are reported in partsper million (ppm, δ) downfield from tetramethylsilane (TMS).Multiplicities of signals in the ¹H NMR spectra are reported as follows:s (singlet), d (doublet), t (triplet), q (quartet), m (multiplet), dd(doublet of doublets), dt (doublet of triplet), etc.

Infrared spectra were obtained on a Perkin Elmer 1600 FT-IR spectrometeras liquid films and thin layer with NaCl cells. Intensities werereported as s (strong 67-100%), in (medium 34-66%), and w (weak 0-33%)with the following notations, hr (broadened), sh (shoulder), etc.

Optical rotations were recorded on JASCO, P-1100 model polarimeter(Japan Spectroscopic Co., Ltd.) with sodium D line at the temperaturesas indicated in the experimental section for the specific compounds.

Analytical Thin Layer Chromatography (TLC) was performed on Merck silicagel plates (Merck Kieselgel, 60, 0.25-mm thickness) with F₂₅₄ indicator.Compounds were visualized under UV lamp and/or by developing withiodine, vanillin, p-anisaldehyde or KMnO₄ followed by heating with aheat gun. Flash chromatography (Still, W. C., et al., J. Org. Chem.,43:1404 (1978)) was performed as reported by Still and coworkers on230-400 mesh silica gel (E.M. Science) with technical and/or HPLC gradesolvents. Medium Pressure Liquid Chromatography (MPLC) was performedwith FMI pump and prepacked silica gel column (Merck, Labor Columns,LiChroprep Si 60, 40-63 μm). High Pressure Liquid Chromatography (HPLC)was performed on a Rainin HPLX system equipped with two 25 mL pump headsand a Rainin Dynamax UV-C dual-beam variable wavelength detector set at254 or 260 nm using Phenomenex, Luna 5μ C18 semipreparative (250×10 mm)column and Chiralcel OJ semipreparative (250×10 mm) column.

Low and high-resolution mass spectra (LRMS and HRMS) were obtained withelectronic or chemical ionization (EI or CI) either (1) at Johns HopkinsUniversity on a VG Instruments 70-S spectrometer run at 70 eV for EI andrun with ammonia (NH₃) as a carrier gas for CI or (2) at the Ohio StateUniversity on a Finnigan-MAT CH5, a Finnigan-MAT 731, or a VGInstruments 70-VSE spectrometer run at 70 eV for EI and run with methane(CH₄) for CI.

Example 1 Synthesis of Compounds 6a-n General Procedure for theSynthesis of Compounds 6a-n.1-[4-(5-Bromopyrimidin-2-ylsulfanyl)phenyl]-3-(2-nitrobenzoyl)-urea (6n)

A solution of 5-bromo-2-chloropyrimidine (5 g, 0.026 mol),4-aminothiophenol (3.24 g, 0.026 mol), and K₂CO₃ (7.14 g, 0.052 mol) indry DMSO (50 mL) was stirred at 120° C. for 2.5 hours under N₂. Aftercooling, the reaction mixture was poured into water and extracted withethyl acetate. The organic layer was washed with water and saturatedbrine and then dried. The solvent was evaporated to give a residue thatwas purified by silica gel flash column chromatography (ethylacetate-hexane 1:3) to give 4-(5-bromopyrimidin-2-ylsulfanyl)phenylamine(4n), yield 74%. ¹H NMR (400 MHz, CDCl₃): δ 8.51 (s, 2H), 7.37 (d, J)8.0 Hz, 2H), 6.72 (d, J) 8.0 Hz, 2H), 2.63 (s, 2H); EI-MS m/z 281 [M]⁺,283 [M+2]⁺.

A solution of 2-nitrobenzoyl isocyanate (3 g, 0.016 mol) in dry1,4-dioxane (15 mL) was added dropwise to a solution of 4n (2.93 g, 0.01mol) in dry 1,4-dioxane (15 mL) with stirring at room temperature. Thereaction mixture was stirred for 18 hours and then diluted with water.The precipitated solid was collected by filtration and washed withwater. The solid was dissolved in ethyl acetate, and the organic layerwas washed with water 2-3 times, dried, and concentrated to give1-[4-(5-bromopyrimidin-2-ylsulfanyl)phenyl]-3-(2-nitrobenzoyl)urea (6n),yield 92%. ¹H NMR (400 MHz, DMSO-d₆): δ 11.32 (br, s, 1H), 10.35 (br, s,1H), 8.78 (s, 2H), 8.22 (m, 1H), 7.91 (m, 1H), 7.77 (m, 2H), 7.67 (m,2H), 7.57 (m 2H); EI-MS m/z 473 [M]⁺, 475 [M+2]⁺; HRMS calculated forC₁₈H₁₂BrN₅O₄SNa [M+Na]⁺: 495.9685, found: 495.9701. Anal.(C₁₈H₁₂BrN₅O₄S) C, H, N.

Example 2 Synthesis of Compounds 7a-d General Procedure for theSynthesis of Compounds 7a-d.1-(2-Aminobenzoyl)-3-[4-(5-bromopyrimidin-2-ylsulfanyl)phenyl]urea (7d)

Iron powder (1.77 g, 31.63 mmol) was added in portions to a mixture of6n (3 g, 6.32 mmol) in AcOH (90 mL) at 80° C. The reaction mixture wasrefluxed for 30 min and then cooled to room temperature and diluted withwater. The precipitated solid was collected by filtration. The solid wasdissolved in an excess of ethyl acetate and filtered. The filtrate wasdried and concentrated to give a residue that was purified by silica gelflash column chromatography (ethyl acetate-hexane 2:3) to give 7d, yield62%. ¹H NMR (400 MHz, DMSO-d₆): δ 11.92 (br, s, 1H), 10.73 (br, s, 1H),8.75 (s, 2H), 7.68 (m, 2H), 7.57 (m, 2H), 7.25 (m, 1H), 6.79 (m, 1H),6.59 (m 2H); EI-MS m/z 443 [M]⁺, 445 [M+2]⁺; HRMS calculated forC₁₈H₁₄BrN₅O₂SNa [M+Na]⁺: 465.9943, found: 465.9938. Anal.(C₁₈H₁₄BrN₅O₂S) C, H, N.

Example 2 Synthesis of Compounds 8a-h General Procedure for theSynthesis of Compounds 8a-h.1-[4-(5-Bromopyrimidine-2-sulfinyl)phenyl]-3-(2-nitrobenzoyl)-urea (8g)

To a stirred solution of 6n (1.0 g, 2.11 mmol) in DCM (50 mL) at 0° C.was added MCPBA (0.364 g, 2.11 mmol) in portions. The resulting mixturewas stirred at room temperature for 6 h and then diluted with water andmade slightly basic with Na₂CO₃ solution. The DCM layer was separated,and the aqueous layer was extracted with DCM. The combined organiclayers were dried and evaporated to give a residue that was purified bysilica gel flash column chromatography (ethyl acetate-methanol 5:1) togive 8g, yield 48%. ¹H NMR (400 MHz, DMSO-d₆): δ 11.32 (br, s, 1H),10.38 (br, s, 1H), 9.15 (s, 2H), 8.20 (m, 1H), 7.88 (m, 1H), 7.72-7.80(m, 6H); EI-MS m/z 489 [M]⁺, 491 [M+2]⁺; HRMS calculated forC₁₈H₁₂BrN₅O₅SNa [M+Na]⁺: 511.9634, found: 511.9632. Anal.(C₁₈H₁₂BrN₅O₅S) C, H, N.

1-[4-(5-Bromopyrimidine-2-sulfonyl)phenyl]-3-(2-nitrobenzoyl)-urea (8h)

The title compound was synthesized from 6n according to above procedureusing 3 equiv of MCPBA, yield 56%. ¹H NMR (400 MHz, DMSO-d₆): δ 11.42(br, s, 1H), 10.55 (br, s, 1H), 9.23 (s, 2H), 8.21 (m, 1H), 7.76-7.97(m, 7H); EI-MS m/z 505 [M]⁺, 507 [M+2]⁺; HRMS calculated forC₁₈H₁₂BrN₅O₆SNa [M+Na]⁺: 527.9583, found: 527.9592. Anal.(C₁₈H₁₂BrN₅O₆S) C, H, N.

The terms and expressions which have been employed herein are used asterms of description and not of limitation, and there is no intention inthe use of such terms and expressions of excluding equivalents of thefeatures shown and described, or portions thereof, it being recognizedthat various modifications are possible within the scope of theinvention claimed. Moreover, any one or more features of any embodimentof the invention may be combined with any one or more other features ofany other embodiment of the invention, without departing from the scopeof the invention. For example, the features of the compounds of thisinvention are equally applicable to the methods of treating diseasestates described herein. All publications, patents, and patentapplications cited herein are hereby incorporated by reference in theirentirety for all purposes.

1. A method of inhibiting tubulin polymerization in a cell, comprisingcontacting said cell with a composition comprising 1 nM to 10 microM ofa compound represented by either of the following formulas:

or a pharmaceutically acceptable salt thereof.
 2. The method of claim 1,wherein said cell is a cancerous cell.
 3. (canceled)
 4. (canceled)
 5. Aprocess for the preparation of1-[4-(5-Bromopyrimidin-2-ylsulfanyl)phenyl]-3-(2-nitrobenzoyl)-ureacomprising the steps of: (a) preparing a solution comprising of5-bromo-2-chloropyrimidine and 4-aminothiophenol and heating saidsolution to yield 4-(5-bromopyrimidin-2-ylsulfanyl)phenylamine; and (b)mixing a solution of said 4-(5-bromopyrimidin-2-ylsulfanyl)phenylamineof step (a) with a solution of 2-nitrobenzoyl isocyanate to yield1-[4-(5-Bromopyrimidin-2-ylsulfanyl)phenyl]-3-(2-nitrobenzoyl)-urea. 6.The process of claim 5, wherein the solvent of said solution of step (a)is a dry or an anhydrous solvent.
 7. The process of claim 6, whereinsaid solvent is a dry dimethylsulfoxide.
 8. (canceled)
 9. The process ofclaim 5, wherein said solution of step (a) further comprises a base. 10.The process of claim 9, wherein said base is potassium carbonate. 11.The process of claim 5, wherein said4-(5-bromopyrimidin-2-ylsulfanyl)phenylamine of step (a) is isolated.12. (canceled)
 13. The process of claim 5, wherein the solvent of saidsolution comprising 4-(5-bromopyrimidin-2-ylsulfanyl)phenylamine of step(b) is a dry or an anhydrous solvent.
 14. The process of claim 13,wherein said solvent is dioxane.
 15. The process of claim 1, wherein thesolvent of said solution of 2-nitrobenzoyl isocyanate of step (b) is adry or an anhydrous solvent.
 16. The process of claim 15, wherein saidsolvent is dioxane.
 17. (canceled)
 18. A process for the preparation of1-(2-Aminobenzoyl)-3-[4-(5-bromopyrimidin-2-ylsulfanyl)phenyl]ureacomprising the steps of: (a) preparing a solution comprising of5-bromo-2-chloropyrimidine and 4-aminothiophenol and heating saidsolution to yield 4-(5-bromopyrimidin-2-ylsulfanyl)phenyl amine; (b)mixing a solution of said 4-(5-bromopyrimidin-2-ylsulfanyl)phenylamineof step (a) with a solution of 2-nitrobenzoyl isocyanate to yield1-[4-(5-Bromopyrimidin-2-ylsulfanyl)phenyl]-3-(2-nitrobenzoyl)-urea; and(c) reduction of1-[4-(5-Bromopyrimidin-2-ylsulfanyl)phenyl]-3-(2-nitrobenzoyl)-urea, toyield1-(2-Aminobenzoyl)-3-[4-(5-bromopyrimidin-2-ylsulfanyl)phenyl]urea. 19.The process of claim 18, wherein the solvent of said solution of step(a) is a dry or an anhydrous solvent.
 20. The process of claim 19,wherein said solvent is dry dimethylsulfoxide.
 21. (canceled)
 22. Theprocess of claim 18, wherein said solution of step (a) further comprisesa base.
 23. The process of claim 22, wherein said base is potassiumcarbonate.
 24. The process of claim 18, wherein said4-(5-bromopyrimidin-2-ylsulfanyl)phenylamine of step (a) is isolated.25. The process of claim 24, wherein said isolated is by extraction withethyl acetate and water.
 26. The process of claim 18, wherein thesolvent of said solution of 4-(5-bromopyrimidin-2-ylsulfanyl)phenylamineof step (b) is a dry or an anhydrous solvent.
 27. The process of claim26, wherein said solvent is dioxane.
 28. The process of claim 18,wherein the solvent of said solution of 2-nitrobenzoyl isocyanate ofstep (b) is a dry or an anhydrous solvent.
 29. The process of claim 28,wherein said solvent is dioxane.
 30. (canceled)
 31. The process of claim18, wherein said reduction is performed using iron (Fe) to obtain the2-NH₂ derivative.